1. Field of the Invention
The present invention relates to a method of designing probes for detecting a target sequence and a method of detecting a target sequence using the probes.
2. Description of the Related Art
Due to recent advances in biotechnology, genomic sequences of many species, including humans, have been identified. Consequently, research on making a microarray for sequence analysis and disease diagnosis has been widely conducted. A microarray comprises a substrate on which a group of polynucleotides are immobilized at high density, wherein each group of polynucleotides is immobilized at fixed locations. A microarray can be used to analyze a target biomolecule to obtain a large amount of biological information (for example, sequence), while requiring a minimal amount of the target biomolecule, such as a nucleic acid or a protein.
For example, when a polynucleotide (also called “a probe”, “a probe nucleic acid”, or “a probe polynucleotide”) is immobilized on the microarray that can be specifically hybridized with a target nucleic acid sequence, the microarray can be used to detect and identify the target nucleic acid sequence.
FIG. 1 is a schematic diagram illustrating an example of a conventional method of designing probes.
Referring to FIG. 1, in this conventional method of designing a probe, one desirable probe sequence is selected that can specifically hybridize with the complement of the shown target sequence, but does not cross-hybridize with other non-target sequences. However, it is difficult to design a specific probe when sequence similarity between the target sequence and non-target sequences is high or when the number of target sequences to be identified by the probe is large.
For example, in order to identify a species of bacteria in a sample comprising a plurality of bacteria, a consensus sequence of the plurality of bacteria, for example, a consensus sequence from 16S rRNA or 23S rRNA, can be used to identify probes specific for the various bacterial species in the sample. Such a method can be used to identify several species of bacteria, but is limited to identification of ten or fewer species of bacteria in a given sample since sequence similarity is remarkably high.
FIG. 2 is a schematic diagram illustrating another example of a conventional method of designing probes.
Referring to FIG. 2, in this second conventional method of designing a probe, all possible probes of a given length which hybridize with the complement of the target sequence shown are selected. The sequences of the probes of a given length are varied by varying the position of the first nucleotide of each probe relative to the target region shown in SEQ ID NO:1. As can be seen in FIG. 2, each probe sequence starts 1 bp further 3′ along SEQ ID NO:1. However, a microarray of probes designed using this method would be expensive to manufacture and would yield results that are difficult and time-consuming to analyze since a large number of probes are used in this method.